Microtubule-Driven Multimerization Recruits ase1p onto Overlapping Microtubules
Microtubule-Driven Multimerization Recruits ase1p onto Overlapping Microtubules
复制标题
DOI：
10.1016/j.cub.2008.09.046
复制
发表时间：
2008-11-11
期刊：
影响因子：
9.2
通讯作者：
Peterman, Erwin J. G.
中科院分区：
文献类型：
--
作者：
Kapitein, Lukas C.;Janson, Marcel E.;Peterman, Erwin J. G.
ase1p/PRC1/Map65家族的微管（MT）交联蛋白在有丝分裂纺锤体等微管网络的构建中发挥重要作用。该家族的大多数同源物已被证明具有显著的特异性定位于具有反平行相对取向重叠的mt集[1-4]。调节蛋白结合到ase1p/PRC1/Map65上，并利用定位建立精确的空间信号[5-10]。在这里，我们提出了一种局部蛋白多聚机制的证据，该机制是基于分裂酵母同源物ase1p的特异性靶向。在体外对照实验中，ase1-GFP二聚体沿单个mt表面扩散，当浓度超过一定阈值时，组装成静态多聚体结构。我们观察到，在重叠的mt上，这个阈值明显较低。我们还观察到活细胞内mt上ase1-GFP的扩散和多聚合，这表明多聚合驱动的定位机制与体内相关。通过一系列的ase1p截断，确定了负责MT结合和多化的结构域。我们的研究结果表明，细胞使用一种精细调整的合作定位机制，利用沿单个和重叠mt的ase1p结合位点的几何和浓度差异。
Microtubule (MT) crosslinking proteins of the ase1p/PRC1/Map65 family play a major role in the construction of MT networks such as the mitotic spindle. Most homologs in this family have been shown to localize with a remarkable specificity to sets of MTs that overlap with an antiparallel relative orientation [1-4]. Regulatory proteins bind to ase1p/PRC1/Map65 and appear to use the localization to set up precise spatial signals [5-10]. Here, we present evidence for a mechanism of localized protein multimerization underlying the specific targeting of ase1p, the fision yeast homolog. In controlled in vitro experiments, dimers of ase1-GFP diffused along the surface of single MTs and, at concentrations above a certain threshold, assembled into static multimeric structures. We observed that this threshold was significantly lower on overlapping MTs. We also observed diffusion and multimerization of ase1-GFP on MTs inside living cells, suggesting that a multimerization-driven localization mechanism is relevant in vivo. The domains responsible for MT binding and multimerization were identified via a series of ase1p truncations. Our findings show that cells use a finely tuned cooperative localization mechanism that exploits differences in the geometry and concentration of ase1p binding sites along single and overlapping MTs.